1. Field of the Invention
This inventions relates to a method and device for the performance of assays that detect molecules; in particular, a method and device for the performance of assays that detect molecules in a liquid medium by virtue of their binding to other molecules.
2. Description of the Prior Art
The association of particular molecules with the health and condition of the body has made it important to develop methods and devices for the detection of molecules that are sensitive, efficient and inexpensive. One approach to such methods and devices is to take advantage of affinities of particular molecules for other molecules. Such affinities have been observed with many types of molecules and are, perhaps, best characterized between nucleic acids (DNA-DNA, DNA-RNA, etc.) and between proteins (enzyme-substrate, receptor-hormone, etc.).
Antibodies are among the most frequently used molecules with affinity for other molecules. The molecules reacting and binding with antibodies are collectively referred to as antigens. Antigens can be of many different types of molecules with many different functions. Prior art uses of antibodies and antigens include both the measurement of antigen with antibody and the measurement of antibody with antigen.
When contemplating the design of a method or device utilizing molecules that react and bind to one another, the primary technical consideration is the means by which bound molecules are separated from unbound reactants. There are a number of approaches to separating bound from unbound reactants known in the prior art.
One approach is to take advantage of the selective absorbing characteristics of a chromatographic matrix. For example, in U.S. Pat. No. 4,205,058, bound and unbound reactants are separated on the basis of size as they passed through a silica gel by capillary action. There are a number of disadvantages to such an approach, however. First, chromatographic matrices that separate molecules by size have molecular dimension requirements necessary for adequate separation. These molecular dimension requirements will change depending on the particular size of the molecules to be separated. Thus, one single matrix will not be adequate for every type of separation. Second, a sizing system must have enough travel distance for the molecules to effect a separation. In the typical case where the matrix is housed in a column, for example, the column must be of sufficient length to allow the unbound reactants to travel away from the bound reactants.
Another approach to separating bound from unbound reactants is to attach and immobilize binding molecules on a solid support so as to create a solid support conjugate. The solid support conjugate with one type of binding molecule is allowed to react with free molecules of a different type. Once the molecule-molecule complex has formed, the mobile, unbound reactants can be separated by exerting a force on the reaction mixture. For example, in U.S. Pat. No. 3,862,303, bound and unbound reactants are separated by using latex beads as a solid support and by centrifugation of the reaction mixture on a density gradient. This method has the disadvantage of the expense and inconvenience of centrifugation equipment.
Another solid support approach is disclosed in U.S. Pat. No. 4,138,474. Bound and unbound reactants are separated in a syringe device by using antibody-containing gel particles as a solid support and by forcing unbound reactants past a barrier through which the gel particles cannot flow. This approach has the disadvantage that the flow rate is controlled by mechanical action and, thus, may not be uniform. Nonuniform flow rates can impact the degree of binding of the reactants and change the sensitivity of the assay.
Still other solid support approaches are disclosed in U.S. Pat. Nos. 4,094,674, 4,168,146, 4,235,601, and 4,366,241. Reactants are separated by using immobilized antibody in an immunoreactive zone as a solid support and by flowing reactants past the immunoreactive zone. These methods have the disadvantage that binding occurs at the time the reactants are separated. Thus, the time allowed for binding is fixed by the flow rate. The amount of time allowed for binding dramatically impacts assay sensitivity.
The present invention presents a solid support approach to the separation of bound and unbound reactants that is sensitive, efficient and inexpensive. The invention does not utilize a sizing system, requires no expensive equipment or mechanical force, and does not separate bound and unbound reactants at the time of binding.
Objects and advantages other than those above set forth will be apparent from the following description when read in connection with the accompanying figures.